Carrimycin ameliorates lipopolysaccharide and cecal ligation and puncture-induced sepsis in mice.

Carrimycin (CA), sanctioned by China's National Medical Products Administration (NMPA) in 2019 for treating acute bronchitis and sinusitis, has recently been observed to exhibit multifaceted biological activities, encompassing anti-inflammatory, antiviral, and anti-tumor properties. Despite these applications, its efficacy in sepsis treatment remains unexplored. This study introduces a novel function of CA, demonstrating its capacity to mitigate sepsis induced by lipopolysaccharide (LPS) and cecal ligation and puncture (CLP) in mice models. Our research employed in vitro assays, real-time quantitative polymerase chain reaction (RT-qPCR), and RNA-seq analysis to establish that CA significantly reduces the levels of pro-inflammatory cytokines, namely tumor necrosis factor-alpha (TNF-α), interleukin 1 beta (IL-1ß), and interleukin 6 (IL-6), in response to LPS stimulation. Additionally, Western blotting and immunofluorescence assays revealed that CA impedes Nuclear Factor Kappa B (NF-κB) activation in LPS-stimulated RAW264.7 cells. Complementing these findings, in vivo experiments demonstrated that CA effectively alleviates LPS- and CLP-triggered organ inflammation in C57BL/6 mice. Further insights were gained through 16S sequencing, highlighting CA's pivotal role in enhancing gut microbiota diversity and modulating metabolic pathways, particularly by augmenting the production of short-chain fatty acids in mice subjected to CLP. Notably, a comparative analysis revealed that CA's anti-inflammatory efficacy surpasses that of equivalent doses of aspirin (ASP) and TIENAM. Collectively, these findings suggest that CA exhibits significant therapeutic potential in sepsis treatment. This discovery provides a foundational theoretical basis for the clinical application of CA in sepsis management.

A Janus adhesive hydrogel sheet for preventing postoperative tissue adhesion of intestinal injuries.

Although adhesive hydrogels represent an alternative to surgical sutures for non-invasive tissue wound sealing, those with indiscriminate adhesion fail to hold wounds while inhibiting postoperative tissue adhesion, thus limiting their application in intestinal repair. In this study, an asymmetric adhesive hydrogel sheet composed mainly of polyacrylic acid (PAA) and gelatin (GA) that can be wet-adhered to the surface of intestinal tissue was developed. One side of the GA-PAA hydrogel sheet was complexed with polyvinyl alcohol (PVA), which shielded the excess adhesion based on a physical barrier. Both sides of the PVA/GA-PAA hydrogel showed distinct adhesive and antiadhesive properties. Intriguingly, the anti-adhesive side showed significant anti-adhesion toward specific proteins. The results of animal experiments showed that the PVA/GA-PAA hydrogel could firmly adhere to the intestine to stop leakage and prevent post-operative tissue adhesion two weeks after surgery. The hematoxylin and eosin (H&E) staining results showed that the damaged intestinal serosa was repaired without tissue adhesion. It is believed that the controllable adhesion of the adhesive hydrogel offers better prospects for intestinal repair.

A drug-delivery depot for epigenetic modulation and enhanced cancer immunotherapy.

DNA methyltransferase inhibitors (DNMTis) have found widespread application in the management of cancer. Zebularine (Zeb), functioning as a demethylating agent, has exhibited notable advantages and enhanced therapeutic efficacy in the realm of tumour immunotherapy. Nevertheless, due to its lack of targeted functionality, standalone Zeb therapy necessitates the administration of a substantially higher dosage. In this investigation, we have devised an innovative nanodrug formulation, comprising the DNA methyltransferase inhibitor Zeb and pH-responsive chitosan (CS), hereinafter referred to as CS-Zeb nanoparticles (NPs). Our findings have unveiled that CS-Zeb NPs manifest heightened drug release within an acidic milieu (pH 5.5) in comparison to a neutral environment (pH 7.4). Furthermore, in vivo studies have conclusively affirmed that, in contrast to equivalent quantities of Zeb in isolation, the nanocomplex significantly curtailed tumour burden and protracted the survival duration of the B16F10 tumour-bearing murine model. Additionally, CS-Zeb NPs elicited an augmentation of CD8+ T cells within the peripheral circulation of mice and tumour-infiltrating lymphocytes (TILs). Notably, the dosage of CS-Zeb NPs was reduced by a remarkable 70-fold when juxtaposed with Zeb administered in isolation. To summarise, our study underscores the potential of CS-Zeb NPs as an alternative chemotherapeutic agent for cancer treatment.

Regulation of IFNß expression: focusing on the role of its promoter and transcription regulators.

IFNß is a single-copy gene without an intron. Under normal circumstances, it shows low or no expression in cells. It is upregulated only when the body needs it or is stimulated. Stimuli bind to the pattern recognition receptors (PRRs) and pass via various signaling pathways to several basic transcriptional regulators, such as IRFs, NF-кB, and AP-1. Subsequently, the transcriptional regulators enter the nucleus and bind to regulatory elements of the IFNß promoter. After various modifications, the position of the nucleosome is altered and the complex is assembled to activate the IFNß expression. However, IFNß regulation involves a complex network. For the study of immunity and diseases, it is important to understand how transcription factors bind to regulatory elements through specific forms, which elements in cells are involved in regulation, what regulation occurs during the assembly of enhancers and transcription complexes, and the possible regulatory mechanisms after transcription. Thus, this review focuses on the various regulatory mechanisms and elements involved in the activation of IFNß expression. In addition, we discuss the impact of this regulation in biology.

Anal fistulotomy with one-stage shaped skin grafting for intersphincter anal fistulas: study protocol on a multicentre randomised controlled trial.

BACKGROUND: Anal fistulas are mainly treated via surgery. They can be difficult to treat without surgical intervention. Numerous procedures, such as fistulectomy and fistulotomy, are performed to treat anal fistulas and achieve good effects. However, the wounds created through fistulectomy and fistulotomy take a long time to heal. Therefore, a multicentre randomised controlled trial (RCT) is proposed to study the efficacy of one-stage shaped skin grafting at the surgical wound to heal low simple intersphincter anal fistulas. METHODS: This study is a multicentre, hospital-based RCT. It will be performed at three hospitals. A total of 104 patients with low simple intersphincter anal fistulas who meet the inclusion criteria will be included in this trial and will be allocated randomly to two groups (test and control groups). The patients in the test group will receive one-stage anal fistulotomy surgery combined with shaped skin grafting, and those in the control group will undergo anal fistulotomy only. All the operations will be performed by attending colorectal surgeons or surgeons of a higher level. Effectiveness and safety indicators will be observed, recorded and analysed. DISCUSSION: Anal fistulotomy can heal low simple intersphincter anal fistulas effectively and safely with a low recurrence rate. Skin grafts promote wound epithelisation significantly. We believe that skin grafting can treat low simple intersphincter fistulas with a short healing time. TRIAL REGISTRATION: Chinese Clinical Trial Register, ChiCTR2000039174. Registered on 28 October 2020.

Doxycycline Attenuates Pig Intestinal Microbial Interactions and Changes Microbial Metabolic Pathways.

Doxycycline is a therapeutic veterinary antibiotic commonly used in pig breeding. In this study, 27 fattening pigs of 33.5 ± 0.72 kg were divided equally into 3 groups. Doxycycline at 0, 3, and 5 mg/kg body weight was added to the feed in groups CK, L and H. The medication and withdrawal periods were set at 5 and 28 days. The results showed that the doxycycline average concentrations in groups L and H during the medication period were 117.63 ± 13.54 and 202.03 ± 24.91 mg/kg dry matter, respectively. Doxycycline levels were lower than the detection limit after 20 days. Doxycycline did not affect the diversity of the intestinal microbial community structure. The relative abundances of Streptococcus were significantly higher in treatment groups than that in group CK, and Alishewanella, Vagococcus, Cloacibacterium, and Campylobacter abundances were significantly positively correlated with doxycycline concentration. Interestingly, the microbiota cooccurrence network suggested that high doxycycline concentration weakened the interactions among bacteria until day 33. Functional prediction showed that doxycycline significantly altered metabolic pathways related to the cell membrane. The results revealed that the use of doxycycline during pig breeding can affect bacterial abundance during the withdrawal period, and it may affect interactions among bacteria and change the intestinal metabolic pathways.

Corrigendum: Changes in the Carbon Metabolism of Escherichia coli During the Evolution of Doxycycline Resistance.

[This corrects the article DOI: 10.3389/fmicb.2019.02506.].

Therapeutic Development by Targeting the cGAS-STING Pathway in Autoimmune Disease and Cancer.

DNA immune recognition regulation mediated by the cGAS-STING pathway plays an important role in immune functions. Under normal physiological conditions, cGAS can recognize and bind to invading pathogen DNA and activate the innate immune response. On the other hand, abnormal activation of cGAS or STING is closely related to autoimmune diseases. In addition, activation of STING proteins as a bridge connecting innate immunity and adaptive immunity can effectively restrain tumor growth. Therefore, targeting the cGAS-STING pathway can alleviate autoimmune symptoms and be a potential drug target for treating cancer. This article summarizes the current progress on cGAS-STING pathway modulators and lays the foundation for further investigating therapeutic development in autoimmune diseases and tumors.

Zebularine elevates STING expression and enhances cGAMP cancer immunotherapy in mice.

DNA methylation abnormality is closely related to tumor occurrence and development. Chemical inhibitors targeting DNA methyltransferase (DNMTis) have been used in treating cancer. However, the impact of DNMTis on antitumor immunity has not been well elucidated. In this study, we show that zebularine (a demethylating agent) treatment of cancer cells led to increased levels of interferon response in a cyclic guanosine monophosphate-AMP (cGAMP) synthase (cGAS)- and stimulator of interferon genes (STING)-dependent manner. This treatment also specifically sensitized the cGAS-STING pathway in response to DNA stimulation. Incorporation of zebularine into genomic DNA caused demethylation and elevated expression of a group of genes, including STING. Without causing DNA damage, zebularine led to accumulation of DNA species in the cytoplasm of treated cells. In syngeneic tumor models, administration of zebularine alone reduced tumor burden and extended mice survival. This effect synergized with cGAMP and immune checkpoint blockade therapy. The efficacy of zebularine was abolished in nude mice and in cGAS-/- or STING-/- mice, indicating its dependency on host immunity. Analysis of tumor cells indicates upregulation of interferon-stimulated genes (ISGs) following zebularine administration. Zebularine promoted infiltration of CD8 T cells and natural killer (NK) cells into tumor and therefore suppressed tumor growth. This study unveils the role of zebularine in sensitizing the cGAS-STING pathway to promote anti-tumor immunity and provides the foundation for further therapeutic development.

Metabonomics reveals an alleviation of fitness cost in resistant E. coli competing against susceptible E. coli at sub-MIC doxycycline.

High concentrations of antibiotics may induce bacterial resistance mutations and further lead to fitness costs by reducing growth of resistant bacteria. However, antibiotic concentrations faced by bacteria are usually low in common environments, which leads to questions about how resistant bacteria with fitness costs regulate metabolism to coexist or compete with susceptible bacteria during sublethal challenge. Our study revealed that a low proportion (< 15%) of resistant bacteria coexisted with susceptible bacteria due to the fitness cost without doxycycline. However, the cost for the resistant strain decreased at a doxycycline concentration of 1 mg/L and even disappeared when the doxycycline concentration was 2 mg/L. Metabonomics analysis revealed that bypass carbon metabolism and biosynthesis of secondary metabolites were the primary metabolic pathways enriching various upregulated metabolites in resistant bacteria without doxycycline. Moreover, the alleviation of fitness cost for resistant bacteria competed with susceptible bacteria at 1 mg/L doxycycline was correlated with the downregulation of the biomarkers pyruvate and pilocarpine. Our study offered new insight into the metabolic mechanisms by which the fitness cost of resistant mutants was reduced at doxycycline concentrations as low as 1 mg/L and identified various potential metabolites to limit the spread of antimicrobial resistance in the environment.

Adding a complex microbial agent twice to the composting of laying-hen manure promoted doxycycline degradation with a low risk on spreading tetracycline resistance genes.

Poultry manure is a reservoir for antibiotics and antibiotic resistance genes and composting is an effective biological treatment for manure. This study explored the effect of using two methods of adding a complex microbial agent to the composting of laying-hen manure on doxycycline degradation and tetracycline resistance genes elimination. The results showed that incorporating a complex microbial agent at 0.8% (w/w) on the 0th and 11th day (group MT2) effectively degraded doxycycline with a final degradation rate of 46.83 ±â€¯0.55%. The half-life of doxycycline in this group was 21.90 ±â€¯0.00 days and was significantly lower than that of group MT1 (1.6% (w/w) complex microbial agent added on the 0th day) and group DT (compost without complex microbial agent). But there was no significant difference in the final degradation rate of doxycycline between group DT and group MT1. The addictive with the complex microbial agent changed the microbial community structure. Bacteroidetes, Firmicutes and Proteobacteria were the dominant phyla during composting. Aerococcus, Desemzia, Facklamia, Lactobacillus, Streptococcus, and Trichococcus were the bacteria related to the degradation of doxycycline. Moreover, the incorporation of a complex microbial agent could decrease the risk on spreading tetracycline resistance genes. The single addition promoted the elimination of tetM, whose possible hosts were Enterococcus, Lactobacillus, Staphylococcus, and Trichococcus. Adding the complex microbial agent twice promoted the elimination of tetX, which was related to the low abundance of Chryseobacterium, Flavobacterium and Neptunomonas in group MT2. Redundancy analysis showed that the bacterial community, residual doxycycline and physiochemical properties have a potential effect on the variation in tetracycline resistance genes levels. Overall, adding the complex microbial agent twice is an effective measure to degrade doxycycline.

Acellular matrix hydrogel for repair of the temporomandibular joint disc.

Application of tissue-derived extracellular matrix (ECM) biomaterials in the repair of the temporomandibular joint (TMJ) disc is a promising approach for the treatment of disc abrasion and perforation, particularly for the young patient population. Although decellularized ECM (dECM) scaffolds preserve tissue-specific structures as well as biological and biomechanical properties, they require surgical implantation. To address this issue, we prepared porcine TMJ discs into decellularized ECM with serial detergent and enzyme treatments, and the TMJ disc-derived ECM was then processed into hydrogels via pepsin digestion. The decellularization efficiency was assessed by quantification of the DNA and matrix component contents. The fibrous ultrastructure of the hydrogel was observed by scanning electron microscopy (SEM). Rheological characterization and mechanical properties were measured. in vitro experiments with costal chondrocytes ensured the cellular proliferative capacity and compatibility in the injectable disc-derived ECM hydrogel. The results showed that a large amount of DNA (>95%) was removed after decellularization; but, the collagen was retained. SEM of the hydrogels demonstrated a multiaperture fiber ultrastructure. Rheological studies revealed a rapid gelation temperature (37°C) and injectable properties. The mechanical properties of the hydrogels were adjusted by changing the ECM concentration. The in vitro studies revealed that the hydrogels are not cytotoxic, but instead showed good cytocompatibility. The hydrogel also showed good injectability and degradability through an in vivo study. Overall, these results suggest the great potential of injectable disc-derived hydrogels for TMJ disc repair and regeneration applications.

Changes in the Carbon Metabolism of Escherichia coli During the Evolution of Doxycycline Resistance.

Despite our continuous improvement in understanding the evolution of antibiotic resistance, the changes in the carbon metabolism during the evolution of antibiotic resistance remains unclear. To investigate the evolution of antibiotic resistance and the changes in carbon metabolism under antibiotic pressure, Escherichia coli K-12 was evolved for 38 passages under a concentration gradient of doxycycline (DOX). The 0th-passage sensitive strain W0, the 20th-passage moderately resistant strain M20, and the 38th-passage highly resistant strain E38 were selected for the determination of biofilm formation, colony area, and carbon metabolism levels, as well as genome and transcriptome sequencing. The MIC of DOX with E. coli significantly increased from 4 to 96 µg/ml, and the IC50 increased from 2.18 ± 0.08 to 64.79 ± 0.75 µg/ml after 38 passages of domestication. Compared with the sensitive strain W0, the biofilm formation amount of the resistant strains M20 and E38 was significantly increased (p < 0.05). Single-nucleotide polymorphisms (SNPs) were distributed in antibiotic resistance-related genes such as ribosome targets, cell membranes, and multiple efflux pumps. In addition, there were no mutated genes related to carbon metabolism. However, the genes involved in the biosynthesis of secondary metabolites and carbon metabolism pathway were downregulated, showing a significant decrease in the metabolic intensity of 23 carbon sources (p < 0.05). The results presented here show that there may be a correlation between the evolution of E. coli DOX resistance and the decrease of carbon metabolism, and the mechanism was worthy of further research, providing a theoretical basis for the prevention and control of microbial resistance.